Synthesis of crosslinked 2′-OMe RNA duplexes and their application for effective inhibition of miRNA function

Abdelhady, Ahmed Mostafa; Hirano, Yu; Onizuka, Kazumitsu; Okamura, Hirokazu; Komatsu, Yasuo; Nagatsugi, Fumi

doi:10.1016/j.bmcl.2021.128257

Cited by 10 publications

(13 citation statements)

References 39 publications

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“…For creating a functional crosslinking structure, we used vinyl chemistry to synthesize crosslinked RNA 27 and 2′-OMe RNA duplexes. 28 This vinyl crosslinking kept a native structure and provided high thermal stability. The native-like crosslinking structure is advantageous for the biochemical study and development of nucleic acid-based medicines because the introduction of large artificial moiety often decreases enzyme binding affinity.…”

Section: Creation Of Interstrand Crosslinking Structuresmentioning

confidence: 99%

Section: Creation Of Interstrand Crosslinking Structuresmentioning

confidence: 99%

Section: Creation Of Interstrand Crosslinking Structuresmentioning

confidence: 99%

“…These crosslinked AMOs containing the antisense-targeting miR-21 showed a markedly higher anti-miRNA activity than the commercially available miR-21 inhibitor, which has locked nucleic acid residues. 28 Prior to the research on AVP-crosslinking AMOs, Komatsu et al reported AMOs with flanking crosslinked-duplex structures, which were prepared using a short bifunctional crosslinker containing bis-aminooxy groups and a pair of apurinic/ apyrimidinic (AP) sites (Fig. 3).…”

Section: Reviewmentioning

confidence: 99%

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Hybridization-specific chemical reactions to create interstrand crosslinking and threaded structures of nucleic acids

et al. 2022

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Section: Creation Of Interstrand Crosslinking Structuresmentioning

confidence: 99%

Section: Creation Of Interstrand Crosslinking Structuresmentioning

confidence: 99%

Section: Creation Of Interstrand Crosslinking Structuresmentioning

confidence: 99%

Section: Reviewmentioning

confidence: 99%

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Hybridization-specific chemical reactions to create interstrand crosslinking and threaded structures of nucleic acids

et al. 2022

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“…The procedures for the microRNA (miRNA) inhibition assay in cells using the AMOs are described in detail. These methods are based on our previously reported work (Abdelhady et al., 2021).…”

Section: Introductionmentioning

confidence: 99%

Synthesis of Crosslinked 2′‐OMe RNA Duplexes Using 2‐Amino‐6‐Vinylpurine and Their Application for Effective Inhibition of miRNA Function

et al. 2022

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Crosslinking reactions to nucleic acids are an effective way to prepare stable complexes formed by covalent bonding. We demonstrated that fully 2′‐O‐methylated (2′‐OMe) RNAs having a 2‐amino‐6‐vinylpurine (AVP) exhibited an efficient crosslinking to uracil in the target RNA. Recently, we reported the preparation of crosslinked 2′‐OMe RNA duplexes using AVP and the anti‐miRNA oligonucleotides (AMOs) containing crosslinked duplexes at the terminal positions. These AMOs exhibited efficient microRNA (miRNA) inhibition at very low concentrations. In this article, we describe the chemical synthesis of 2′‐OMe oligonucleotides containing AVP and preparation of the AMOs bearing crosslinked 2′‐OMe RNA duplexes using AVP. In addition, we describe in detail the miRNA inhibition assay using these AMOs. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Synthesis of phosphoramidite of 2‐amino‐6‐vinylguanosine derivative Basic Protocol 2: Synthesis of AVP‐2′‐OMe RNA Basic Protocol 3: Evaluation of the crosslink reactivity of CFO containing AVP to the 2′‐OMe RNA and preparation of AMOs containing crosslinked duplex Basic Protocol 4: miRNA inhibition assays

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Uracil‐Selective Cross‐Linking in RNA and Inhibition of miRNA Function by 2‐Amino‐6‐vinyl‐7‐deazapurine Deoxynucleosides

Soemawisastra,

Okamura,

Abdelhady

et al. 2024

ChemBioChem

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MicroRNAs (miRNAs) regulate gene expression through RNA interference. Consequently, miRNA inhibitors, such as anti‐miRNA oligonucleotides (AMOs), have attracted attention for treating miRNA overexpression. To achieve efficient inhibition, we developed 2‐amino‐6‐vinylpurine (AVP) nucleosides that form covalent bonds with uridine counterparts in RNA. We demonstrated that mRNA cross‐linked with AVP‐conjugated antisense oligonucleotides with AVP were protected from gene silencing by exogenous miRNA. However, endogenous miRNA function could not be inhibited in cells, probably because of slow cross‐linking kinetics. We recently developed ADpVP, an AVP derivative bearing a 7‐propynyl group—which boasts faster reaction rate than the original AVP. Here, we synthesized dADpVP—a deoxy analog of ADpVP—through a simplified synthesis protocol. Evaluation of the cross‐linking reaction revealed that the reaction kinetics of dADpVP were comparable to those of ADpVP. In addition, structural analysis of the cross‐linked adduct discovered N3 linkage against uridine. Incorporating dADpVP into two types of miRNA inhibitors revealed a marginal impact on AMO efficacy yet improved the performance of target site blockers. These results indicate the potential of cross‐linking nucleosides for indirect miRNA function inhibition.

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Synthesis of crosslinked 2′-OMe RNA duplexes and their application for effective inhibition of miRNA function

Cited by 10 publications

References 39 publications

Hybridization-specific chemical reactions to create interstrand crosslinking and threaded structures of nucleic acids

Hybridization-specific chemical reactions to create interstrand crosslinking and threaded structures of nucleic acids

Synthesis of Crosslinked 2′‐OMe RNA Duplexes Using 2‐Amino‐6‐Vinylpurine and Their Application for Effective Inhibition of miRNA Function

Uracil‐Selective Cross‐Linking in RNA and Inhibition of miRNA Function by 2‐Amino‐6‐vinyl‐7‐deazapurine Deoxynucleosides

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